The long term commercial objective of the proposed research is to develop, formulate and commercialize a growth factors-based, feeder-free culture system for the ex vivo expansion and maintenance of human embryonic stem (hES) cells. Maintenance of undifferentiated hES cells currently requires culture on mouse embryonic fibroblast (MEF) feeders, which represents a major burden in promoting the clinical application of hES cells. A feeder-free culture system for hES cells expansion is an essential technology for using stem-cell therapy in the treatment or management of a disease, condition, or injury.Specific aims during Phase I will include: 1. usage of cytokine expression array to identify potential cytokines involved in the function of MEF feeders to support hES cell growth. 2. investigation of cytokines identified for their functional significance in supporting hES cell culture. Human ES cell line H9 (NIH code WAO9) (WiCeII Research Institute, Wisconsin) will be used for this part of study. Functional significance of these factors can be evaluated by the effect of depletion of individual factor from MEF-conditioned media on hES cell culture.During phase II, the primary growth factors identified during Phase I will be further screened, characterized and used as a base recipe to develop, formulate and commercialize a growth factors-based, feeder-free ex vivo culture system for hES cells.